Until recently isolation and/or purification of nucleic acids from complex mixtures as described above was a laborious, multi-step procedure. In EP 0389063, incorporated herein by reference, a simple and rapid purification of nucleic acid material from a complex mixture is disclosed. This procedure comprises treating the complex mixture, such as whole blood with a chaotropic agent in the presence of a nucleic acid binding silica solid phase material under conditions that allow for binding of all nucleic acid material to said solid phase and separating said solid phase from the mixture. The reference shows that both single stranded and double stranded nucleic acids are bound to the solid phase if present in a mixture. The reference also discloses amplification (PCR) of a certain nucleic acid with a known sequence, suspected to be present in a mixture.
Thus, said reference teaches a simple and rapid detection method for known nucleic acids suspected to be present in a sample.
In many cases the nature of the target nucleic acid (double, stranded or single stranded) may not be known beforehand, or there may be many different targets necessary to be analyzed. In these cases the rapid but rather crude method described above may not be sophisticated enough and further separations of the crude material may be desired. Fractionation of mixtures of double-(ds) and single-stranded (ss) nucleic acids (NA) into single- and double-stranded forms is frequently needed e.g. in the separation of labelled ss-NA probes from ds-hybrids, in the separation of in vitro transcripts from ds-DNA templates, and in the separation of genomic DNA from mRNA. Currently, the separation of different kinds of nucleic acids can be accomplished by several techniques. Electrophoresis can be used to fractionate different forms of nucleic acids, because of differences in size and shape (1–3). Centrifugation takes advantage of differences in density (4), and more recently the technology of high-performence liquid chromatography (HPLC) has been applied to separate and purify single- and double-stranded DNA and RNA molecules (5–8).
RNA purified from eukaryotic cells by the currently most widely used procedure (9) appears to contain significant amounts of genomic DNA. An adaptation which reduces genomic DNA contamination of the ss-RNA fraction has recently been described (10).
It is not possible to look at single stranded and/or double stranded material separately using the method of EP 0389063 because the method does not discriminate between the two.